CN106885906B - The microdose urine protein reagent and detection method of a kind of stabilization, strong antijamming capability - Google Patents
The microdose urine protein reagent and detection method of a kind of stabilization, strong antijamming capability Download PDFInfo
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- CN106885906B CN106885906B CN201510942727.6A CN201510942727A CN106885906B CN 106885906 B CN106885906 B CN 106885906B CN 201510942727 A CN201510942727 A CN 201510942727A CN 106885906 B CN106885906 B CN 106885906B
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- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6803—General methods of protein analysis not limited to specific proteins or families of proteins
- G01N33/6827—Total protein determination, e.g. albumin in urine
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Abstract
The present invention relates to microdose urine protein detection technique fields, more particularly to a kind of cardiac muscle troponin I detection reagent contains 50 mmol/L pH, 7.6 Mes buffer solutions, 25 ml/L TritonX 100 in reagent R1,30 ml/L latex particles, 7.7 mmol/L Sodium azides；Contain 50 mmol/L pH, 7.6 Mes buffer solutions, 30ml/L sheep anti-human albumin antibodies, 20 mmol/L mannitol, 7.7 mmol/L Sodium azides in reagent R2.Using Mes fliud flushings, latex particle and mannitol alcohol are added, the anti-interference ability and stability of reagent are significantly improved, is a kind of more stable, strong antijamming capability microdose urine protein reagent.
Description
Technical field
The present invention relates to microdose urine protein detection technique field, more particularly to a kind of microdose urine protein detection reagent,
Further relate to the detection method using this detection reagent.
Background technology
The molecular weight of microdose urine protein be 6.9 kD, it is negatively charged, be glomerular filtration membrane can by minimum egg
One of white matter molecule accounts for the 40% of filtration Tot Prot.Protein almost all through glomerular filtration is by proximal convoluted tubule with actively
Mode resorption is received, therefore there was only microalbumin in urinating.The discharge rate of microdose urine protein is by the position of people, movement, blood pressure, egg
The influence of the factors such as white intake.Fever, stress can be such that microdose urine protein secretion increases；Some drugs can pass through
Biological impact makes microdose urine protein discharge increase.Under normal circumstances, most albumen cannot be by filtration membrane, but in disease
In the case of reason, such as various inflammation, metabolic disorder and immunologic mjury keeps glomerular hemodynamics abnormal, glomerular filtration membrane damage
Evil is to cause the increased major reason of microdose urine protein discharge rate.Increase since long term hyperglycemia makes non-enzymatic sugar be acylated rate,
Anoxic, blood viscosity is caused to increase, while the endothelial cell release vaso-active substances such as Endothelin and nitric oxide make glomerulus
Capillary tension change, and then cause renal hemodynamics to change, glomerulus is made to be in high filtration state and lead to renal damage.
Early detection microdose urine protein is to preventing and delaying the development of diabetic nephropathy from having highly important clinical meaning.
Microdose urine protein refer to urine in albumin increase, and urinate total protein in term of reference or test-paper method be the moon
Property result.The important indicator that its detection is diagnosed as Renal Injury is in widespread attention, it is to diabetes early stage diagnosis and treatment
And its prognosis is of great significance.It is now recognized that microdose urine protein is the early stage external index of diabetic nephropathy, the present invention adopts
Use Immunity transmission turbidity.
Invention content
It is urinated the object of the present invention is to provide a kind of reagent for detecting microdose urine protein and using reagent detection micro-
The method for measuring albumin content.The kit specifically binds method using Ag-Ab, can effectively detect microdose urine protein
Content, have many advantages, such as that strong antijamming capability, stability are good.
Basic principle：
This method is a kind of Immunity transmission turbidity.Specific antibody is incorporated into latex particle surface, sample and latex
Particle mixes in Mes buffer solutions, and the microdose urine protein in sample is combined with the antibody on latex particle surface, makes adjacent glue
Newborn particle is cross-linked to each other, and solution turbidity variation is detected under 415nm wavelength, and variation degree contains with the microdose urine protein in sample
It measures directly proportional.
What the present invention was obtained through the following steps：
A kind of microdose urine protein detection reagent, including reagent R1 and reagent R2, the composition of the reagent R1 and reagent R2
It is as follows：
1）The group of reagent R1 is divided into：
Mes pH of buffer 7.6······························50 mmol/L；
TritonX-100···································25 ml/L；
Latex particle······································30 ml/L；
Sodium azide········································7.7 mmol/L；
2）The group of reagent R2 is divided into：
Mes pH of buffer 7.6······························50 mmol/L；
Sheep anti-human albumin antibodies······························3 g/L；
Mannitol········································20 mmol/L；
Sodium azide········································7.7 mmol/L。
The microdose urine protein detection reagent detects the detection method of microdose urine protein, uses full-automatic biochemical
Analyzer Hitachi 7180, is measured using end-point method, and detection dominant wavelength is 415 nm.
The ratio of the detection method, R1 reagents and R2 reagents is 3:1.
Beneficial effects of the present invention：
1）Using Mes buffer systems, the stability of reagent is significantly improved；
2）Using latex particle and mannitol, the performance of measurement is not only significantly improved, but also enhances the stability of reagent
And anti-interference ability；
3）It the accuracy of reagent and has good stability, it is cheap, it is easy to use, it can meet clinical needs completely.
Description of the drawings
Fig. 1 is the correlation curve figure of two kinds of reagents,
Fig. 2 is that two kinds of reagents imitate phase stability curve figure.
Specific implementation mode
With reference to specific embodiment, invention is further explained：
Embodiment 1
The detection reagent of microdose urine protein, including reagent R1 and reagent R2：
1）The group of reagent R1 is divided into：
Mes pH of buffer 7.6··································50 mmol/L；
TritonX-100·······································25 ml/L；
Latex particle··········································30 ml/L；
Sodium azide············································7.7 mmol/L；
2) group of reagent R2 is divided into：
Mes pH of buffer 7.6··································50 mmol/L；
Sheep anti-human albumin antibodies··································3 g/L；
Mannitol············································20 mmol/L；
Sodium azide····························· · ··············7.7 mmol/L；
3）The application method of the present embodiment reagent：
The microdose urine protein detection reagent of the present embodiment description, when in use using full-automatic with double reagent function
Biochemical Analyzer, such as 7180 fully-automatic analyzer of Hitachi, are measured using end-point method.By R1 and R2 according to 5:1 ratio
It is placed on corresponding reagent position, distilled water, standard items and sample, operation such as table 1 is placed in the corresponding position of sample disc.
1 embodiment of table, 1 reagent test method
It calculates：Microdose urine protein content（mg/L）=（A measures ÷ A standards）× C standards.
Embodiment 2
Interference is tested：Fresh mix urine is taken, 2 equal portions are divided into, 5 equal portions then will be separated into per equal portions, is added different
Interfering substance, so that its concentration in urine is reached the requirement of table 2.Then 1 gained reagent of embodiment is used respectively, it is normal with market
See and the microdose urine protein reagent approved simultaneously in comparative determination urine microdose urine protein content, control group measurement result
It is shown in Table 2 with the measurement result of each group after addition disturbance substance.Relative deviation (%)=(the measurement mean value-of interference sample
The measurement mean value of check sample)/check sample measurement mean value × 100%.
As can be seen from Table 2,1 reagent of embodiment is in ascorbic acid≤27.3 μm ol/L, uric acid≤240 μm ol/L, urine
Element≤6.5 the U/L of mmol/L, NAG≤9 test result is not significantly interfered with；And group reagent is compareed in above-mentioned concentration chaff interferent
It in the presence of matter, is significantly interfered with, this illustrates that the interference free performance of 1 reagent of embodiment is far superior to contrast agent.
2 embodiment reagent interference free performance of table compares
Embodiment 3
Correlation is tested：It is formulated reagent preparation, the common State Food and Drug Administration with market using embodiment 1
The microdose urine protein kit for certain company approved carries out control test, while having detected 20 clinical urine specimens, detection
The results are shown in Table 3.And obtain the correlation curve of two kinds of reagents（As shown in Figure 1）, shown by testing result, two examinations
The related coefficient of agent box is 0.999, illustrates that the two has great correlation.
3 embodiment of table, 1 reagent and market is common and the cardiac muscle troponin I assay kit contrasting detection knot that gets the nod
Fruit
Embodiment 4
The stability contrast of reagent is tested：To the reagent in embodiment 1, uniformly 13 groups of packing, every group of amount of reagent is R1
It is 5 mL for 25 mL, R2；And the urine for certain company that the State Food and Drug Administration for taking 13 groups of markets common is approved is micro-
Amount albumin reagent box compares.It is placed into 2-8 DEG C of refrigerator, one group of 1 reagent of embodiment of taking-up on the same day monthly and control
Group reagent detects same mixing urine specimen, testing result as shown in Fig. 2, 1 reagent of embodiment in the 2-8 DEG C of city condition of storage Xia Bi
The common microdose urine protein assay kit in field is more stablized.
By verification, this reagent and similar detection reagent comparison correlation are good, and clinical detection sample results are consistent, Neng Gouda
To market to the application requirement of product, and good in anti-interference performance, be it is a kind of more stablize, the detection of good microdose urine protein
Reagent.
Claims (1)
1. a kind of microdose urine protein detection reagent, it is characterised in that including reagent R1 and reagent R2, the reagent R1 and described
The composition of reagent R2 is as follows：
1) group of reagent R1 is divided into：
Mes pH of buffer 7.650mmol/L；
TritonX-100········25ml/L；
Latex particle 30ml/L；
Sodium azide 7.7mmol/L；
2) group of reagent R2 is divided into：
Mes pH of buffer 7.650mmol/L；
Sheep anti-human albumin antibodies 3g/L；
Mannitol 20mmol/L；
Sodium azide 7.7mmol/L；
The ratio of the R1 reagents and the R2 reagents is 5:1.
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